Human embryonic stem cells (hESCs) are pluripotent cells are able to differentiate into all derivatives of the three primary germ layers: ectoderm, endoderm, and mesoderm. This video describes how to prepare cryopreserved hESCs for long-term storage.
A plasmid DNA extraction is a method used to extract and purify plasmid DNA. Kits are available from varying manufacturers, which are named by size of bacterial culture. Megaprep is used when the starting E. coli culture volume is 500 ml~2.5 L of LB broth and the expected DNA yield is 1.5~2.5 mg.
It can be used after a transformation to screen colonies for the plasmid. Primers designed for the insert sequence should be used when preparing the PCR reaction. Thus, any colonies which give rise to an amplification product are likely to contain the correct DNA sequence.
Pipettes are the laboratory instruments used to transport a measured volume of liquid. This video shows you how to wash the reusable glass pipettes.
Protein can be directly quantitated using UV absorbance (280nm). 280nm Absorbance depends on the Tyr and Trp (amino acids with aromatic rings) content. A very rough protein concentration can be obtained by making the assumption that the protein sample has an extinction coefficient of 1 (1 OD = 1 mg/ml protein).
Automated protein synthesizer is an expert system in transcription, translation, and purification of recombinant protein. It is based on the eukaryotic translation apparatus of wheat germ. This high throughput system produces recombinant proteins in vitro. The generated proteins display foldings and biological functions similar to the mammalian counterparts.
Protein can be directly quantitated by colorimetric. Coomassie Blue dye binds to peptides in an acidic medium. The color change of this assay is measured at 595 or 600 nm. We calculate the protein concentration using the formula created by standards.
Protemist® XE is a fully-automated desktop protein synthesizer for large scale protein production using the wheat germ cell-free expression system. It is ideally suited for producing 10 mg to over 500 mg of protein for structural analysis, functional analysis, and assay development.
Following PCR, you need to get rid of excess short primers, dNTPs, enzymes, short-failed PCR products and salts. We use a silica-gel-membrane for binding of DNA in high-salt buffer (pH 4.5~5.5) and elution of DNA with low-salt buffer (pH 7.0~9.0) or ddH2O to get clean PCR products for downstream applications.
The bicinchoninic acid assay (BCA Assay) is a biochemical assay for determining the total level of protein in a solution, similar to Bradford protein assay. The total protein concentration is exhibited by a color change of the sample solution from green to purple in proportion to protein concentration, which can then be measured using colorimetric techniques.
Bradford assay is a rapid and accurate method to determine the concentration of protein. The assay is based on the observation that the absorbance maximum for an acidic solution of Coomassie Brilliant Blue G-250 shifts from 465 nm to 595 nm when binding to protein occurs. Both hydrophobic and ionic interactions stabilize the anionic form of the dye, causing a visible color change. Within the linear range of the assay (~5-25 mg/ml), the more protein present, the more Coomassie binds.
Silver staining is used to detect proteins after electrophoretic separation on polyacrylamide gels. It combines excellent sensitivity while using very simple and cheap equipment and chemicals. The procedures of silver staining are protein fixation, silver impregnation and image development.
A plasmid DNA extraction is a method used to extract and purify plasmid DNA. Kits are available from varying manufacturers, which are named by size of bacterial culture. Gigaprep is used when the starting E. coli culture volume is 2.5~5 L of LB broth and the expected DNA yield is 7.5~10 mg.
The purpose of transformation is to introduce a foreign plasmid into bacteria and the bacteria will replicate the foreign plasmid along with their own DNA. Bacteria which are able to uptake DNA after a heat shock are called "competent".
Propidium iodide (or PI) is an intercalating agent and a fluorescent molecule that can be used to stain DNA. PI is used as a DNA stain for both flow cytometry to evaluate cell viability or DNA content in cell cycle analysis and microscopy to visualize the nucleus and other DNA containing organelles. It can be used to differentiate necrotic, apoptotic and normal cells.
Protemist® DT II is a new generation, fully automated, robotic desktop protein synthesize. It synthesizes protein of interest using wheat germ cell-free system and through bi-layer reaction. We show you the reagents preparation and how to operate the machine step by step.
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Protemist® DT II is a new generation, fully automated, robotic desktop protein synthesize. It synthesizes protein of interest using wheat germ cell-free system and through bi-layer reaction. ProtemistR DT II can run two scales of transcription and translation reactions; either 1.2 ml or 6 ml. It is an expert system in transcription, translation, and purification.
PCR can be used to make changes to the nucleotide sequence of DNA. This is called PCR mutagenesis. Site-direct mutagenesis is one kind of PCR mutagenesis, which introduces a mutation at a specific location on the DNA strand.
Digestion is the process of cutting DNA molecules with restriction endonucleases. These special enzymes recognize specific sequences in the DNA molecule (for example GATATC) wherever that sequence occurs in the DNA. In PCR cloning, insert and vector are cutted by the same restriction endonucleases and then join together in the next step, ligation.